Blood transfusions are linked to the development of acute lung injury and post-injury multiple organ failure (ALI/MOF). Epidemiological studies have associated infusion of older stored packed red blood cells (PRBCs) to the development of ALI/MOF. Acute organ injury, as demonstrated in both animal and in vitro models, is the result of at least two clinical events. The first event causes endothelial cell (EC) activation as evidenced by an increase in adhesion molecules and the synthesis and extracellular release of chemokines, resulting in PMN priming and adherence. Priming not only maximizes the release of both oxidative and nonoxidative cytotoxic agents that comprise the microbicidal arsenal of the PMN, but priming also changes the PMN phenotype to "hyper-responsive", such that stimuli that do not activate resting PMNs activate primed PMNs. The second event causes activation of these adherent "hyper-responsive" PMNs culminating in endothelial damage, capillary leak, and organ injury. Lipids accumulate during the routine storage of blood, and preliminary data (ours) have implicated these compounds in the activation of human pulmonary microvascular endothelial cells (HMVECs). Thus, because EC activation is a requirement for ALI/MOF and its abrogation inhibits ALI/MOF in vitro and in vivo, we hypothesize that the neutral lipophilic compounds in stored blood predispose injured patients to post-injury ALI/MOF by alteration of normal PMN: EC physiology. This alteration causes indiscriminant EC activation resulting in sequestration of hyper-reactive PMNs in specific organ beds that leads to post-injury ALI/MOF. This hypothesis will be tested by completion of the following specific aims. 1: To identify the lipids that accumulate during routine blood storage and are infused during resuscitation in both PRBCs and LRPRBCs. 2: To delineate the signaling pathways activated by these lipophilic compounds that cause altered PMN: EC physiology by examining PMNs and primary ECs, HMVECs and human liver sinusoidal ECs. 3: To test the ability of these lipids to produce ALI as the first event in a two-event, PMN- mediated model of ALI. 4. To elucidate methods to inhibit the accumulation of these compounds during routine storage of both PRBCs and LR-PRBCs. Completion of these specific aims will likely result in methods to abrogate or to ameliorate the clinical effects of these lipids and will ultimately make transfusions safer.